Control of expression of eucaryotic genes is likely to result at least in part from sequence elements which dictate the precise site of transcription initiation and the frequency of these initiation events. Superimposed upon the sequences which define the eucaryotic promoter is a complex hierarchy of controls likely to involve regulatory molecules dictating the level of transcription in specific tissues and specific stages in development or in response to different external stimuli. In this research, we address two regulated systems in the mammalian cells: the expression of human growth hormone RNA by the pituitary and the expression of globin RNA by the erythroid cells. In preliminary studies, we have utilized in vitro mutagenesis in concert with subsequent DNA transfer experiments to identify DNA sequences 5' to the structural genes for growth hormone and globin which render these genes responsive to specific inducers. These experiments ask: What is the number, sequence and location of regulatory sequences about these genes which render them responsive to induction? Can we identify the nature of the regulatory molecules and clone the genes encoding them? And finally, how do regulatory molecules interact with specific sequences about inducible genes to generate a productive complex enhancing the frequency of transcription? To understand the parameters controlling gene expression, it may be essential to insert wild-type and mutant genes into cells in their original endogenous location within the chromosome. We therefore describe the development of gene transfer procedures in the mammalian cells which may permit homologous recombination. The ability to introduce specific wild-type genes in vitro constructed mutants into cells under conditions in which they function appropriately affords the unique opportunity to discern those sequences essential for the various levels of expression. Further, this system may provide essential technologies required for the development of successful gene therapy.